Application of Proteonano™ Plasma Proteome Enrich Kit

Proteonano™ Technical Principle

The Proteonano™ technology utilizes magnetic nanoparticles (multivalent, multi-affinity nano-probes) whose surfaces are modified with peptides to capture proteins in biofluid samples, enabling selective capture and enrichment of low-abundant proteins. The proprietary designed peptides are chemically coupled to superparamagnetic nanoparticles with a diameter of only nm and absolute homogeneity. Each nanoparticle has approximately 300,000 protein binding sites, allowing it to bind and enrich thousands of proteins in a very small space. In addition, each nanoparticle has a volume of only 10 µm³, which is one billionth of the volume of traditional immunological methods. The extremely small volume increases the sensitivity of detecting low-concentration proteins by 1 million times.

Therefore, the Proteonano™ technology can enrich low-abundant proteins in biological samples, enabling unprecedented depth and throughput of proteomic analysis. This makes Proteonano™ technology a powerful solution for discovering large-scale protein biomarkers.

Proteonano™ Plasma Proteome Enrich Kit

Amerigo Scientific offers the Proteonano™ plasma proteome enrich kit, which is an efficient and user-friendly tool for enriching low-abundance proteins.

Product Features

• This kit has strong compatibility, and it can be compatible with the Nanomation G1 Series and other third-party automated sample processing systems.
• The Proteonano™ plasma proteome enrich kit contains nanoparticles for enrichment, pre-treatment reagents, rapid trypsin, and desalting column.
• The required sample volume is only 20 microliters of plasma/serum, enabling high stability and reproducibility with a median CV of less than 15%.

Experimental Procedures

Fig. 1 Experimental procedures of proteonano™ plasma proteome enrich kit

Comparison of Plasma Enrichment Kits

The plasma enrichment kits available on the market are usually designed to enrich low-abundance proteins from complex samples. Biofluid samples are directly used as inputs, which are then processed into peptides for liquid chromatography-mass spectrometry analysis.

Identification of Biomarkers for Alzheimer's Disease

In the identification of biomarkers for Alzheimer's disease, over 4,000 proteins were identified from plasma samples using the Proteonano™ plasma proteome enrich kit. 159 significantly different proteins were analyzed to be associated with the progression of cognitive impairment, and a multivariate model was constructed with an AUC value reaching 0.92.

Protein Groups (PGs) Identified

4347 PGs were identified in 206 plasma samples. 2704 of them were mapped to the Human Plasma Proteome Project (HPPP) protein catalog. Concentrations of these proteins spanned 9 orders of magnitude, with lowest protein concentration of 7.6 pg/ml.

Figure 2. Protein groups identified in samples

Figure 3. CV% of identified protein groups

Figure 4. Estimated plasma concentrations (ng/mL) of identified proteins

Differential Protein Analysis

The differential protein analysis showed 64, 97, and 159 proteins had different abundance between MCI and N, D and MCI, and D and N groups, respectively. The most upregulated protein in MCI group relative to N group was non-erythrocytic β spectrin (SPTBN1) and most downregulated protein was matrix Gla protein (MGP).

Figure 5. Differential protein analysis

Figure 6. True positive versus false positive rate

Multivariate Analysis

In the multivariate analysis, eight feature selection methods were employed, such as the Least Absolute Shrinkage and Selection Operator (LASSO) and Random Forest (RF). The best model for differentiating the MCI group from the N group contains nine proteins, with the ROC-AUC value of 0.92 (5-95% confidence interval: 0.89-0.98) and the PR-AUC value of 0.84.

Figure 7. Feature selection methods for multivariate analysis